The present invention relates to a process for the production of erythropoietin and, more particularly, to a method for the preparation of partially purified erythropoietin by separating from human urine by means of adsorption.
Erythropoietin is a hormone specifically stimulating erythropoiesis. It is an essential factor for the differentiation of hematopoietic stem cells into matured erythrocytes so that deficiency thereof causes anemia. Therefore, erythropoietin is a promising medicine for curing anemia or, in particular, renal anemia but unfortunately it is not in practical use due to its low availability.
Erythropoietin is a kind of protein so that, when it is intended to be used as a medicine, it is desirably to be prepared from a raw material of human origin in consideration of the possible antigenicity. As the starting raw material of human origin for the preparation of erythropoietin, there may be proposed, for example, human blood or urine from patients suffering from aplastic anemia or the like desease who excrete large amounts of erythropoietin in their urine. These raw materials are, however, limited in the quantity of supply. Accordingly, it will be the most advantageous way to develop a practical method for the preparation thereof from healthy human urine available in large volumes since much larger amounts can be expected as a total content of erythropoietin despite the low concentration thereof than with the above mentioned raw materials.
There have been hitherto made several attempts to obtain purified erythropoietin from human-origin raw materials. These attempts, however, are directed to the separation of erythropoietin from raw materials in small volumes, such as human blood or urine of anemic patients. These prior art methods are not suitable for treating large volumes of healthy human urine containing erythropoietin only in a low concentration with respect to the degree of concentration, yield of recovery, operability of the process and the production costs. A promising and practical way for separating erythropoietin from a large volume of human urine is an adsorption method with an adsorbent which is directly contacted with urine. No method in this way of direct adsorption has been proposed hitherto for treating not only urine from healthy human but also urine from anemic patients.
All of the adsorption methods hitherto proposed are preceded by a preliminary concentration or partial purification of erythropoietin by means of precipitation with a suitable precipitant or an organic solvent, salting out, dialysis and the like methods and the thus partially purified erythropoietin is adsorbed on an adsorbent such as, for example, anionic ion-exchange materials. The inventors have tried direct adsorption of erythropoietin on an adsorbent such as diethylaminoethylcellulose, strongly or weakly basic anionic ion-exchange resins and the like by directly contacting these adsorbents with human urine but without success since erythropoietin is scarcely adsorbed on these adsorbents.